Research on patients with advanced esophageal squamous cell carcinoma (ESCC) who have progressed on immunotherapy remains limited. BL-B01D1 is a first-in-class antibody-drug conjugate consisting of an EGFR-HER3 bispecific antibody bound to a topoisomerase I inhibitor (Ed-04) payload via a cleavable linker. Here, we present safety and efficacy data from a phase 1 study of BL-B01D1, in 82 patients previously treated for ESCC. The primary endpoint was the recommended phase 2 dose. Administered doses were 2.0 (n = 22) and 2.5 (n = 60) mg kg-1 D1D8 infusion every 3 weeks (Q3W). The confirmed objective response rate (cORR) was 29.3% (24 of 82) in all patients and 32.9% (24 of 73) among evaluable patients. For patients dosed at 2.5 mg kg-1, cORR was 39.6% (21 of 53) and disease control rate was 79.2% (42 of 53). In the 2.0 mg kg-1 group, cORR was 15.0% (3 of 20), and the disease control rate was 50.0% (10 of 20). The phase 2 dose was established at 2.5 mg kg-1 D1D8 Q3W. The incidence of G3 treatment-related adverse events at 2.5 mg kg-1 was 63.3%; most common adverse events were anemia (28.3%), leukopenia and thrombocytopenia (18.3%, each), and neutropenia (16.7%). Two cases of ≥G3 interstitial lung disease were observed. Overall, BL-B01D1 demonstrated promising efficacy and manageable safety in patients with metastatic ESCC. A further phase 3 clinical trial has already been initiated. ClinicalTrials.gov registration: NCT05262491 .© 2025. The Author(s).